SELECTED ISOZYMES OF PKC CONTRIBUTE TO AUGMENTED GROWTH OF FETAL AND NEONATAL BOVINE PA ADVENTITIAL FIBROBLASTS

We sought to determine which isozymes of protein kinase C (PKC) contri bute to the increased proliferation of immature bovine pulmonary arter y (PA) adventitial fibroblasts. Seven were identified in lysates of ne onatal PA fibroblasts by Western blot: three Ca2+ dependent (alpha, be ta I, and beta II) and four Ca2+ independent (delta, epsilon, zeta, an d mu). Four isozymes (gamma, eta, theta, and iota) were not detected i n fibroblasts isolated at any developmental stage. Of the seven detect ed isozymes, only PKC-alpha and -beta II protein levels were higher in fetal and neonatal cells compared with adult fibroblasts. Their role in the enhanced growth of immature fibroblasts was then evaluated. The isozyme nonselective PKC inhibitor Ro-31-8220 was first compared with GF-109203X, a structural analog of Ro-31-8220 with relative specifici ty for the Ca2+-dependent isozymes of PKC. GF-109203X selectively inhi bited the growth of immature cells and was nearly as potent as Ro-31-8 220. Go-6976, a more specific inhibitor of the Ca2+-dependent isozymes , mimicked the antiproliferative effect of GF-109203X. PKC downregulat ion with 1 mu M phorbol 12-myristate 13-acetate had the same selective antiproliferative effect on immature fibroblasts as GF-109203X and Go -6976. The protein levels of PKC-alpha and -beta II, but not of PKC-be ta I, were completely degraded in response to phorbol 12-myristate 13- acetate pretreatment. These results suggest that PKC-alpha and -beta I I are important in the augmented growth of immature bovine PA adventit ial fibroblasts.